*Target (protein/gene name): DNA-directed RNA polymerase III subunit *NCBI Gene # or RefSeq#: 5473365 *Protein ID (NP or XP #) or Wolbachia#: XP_001614080.1 *Organism (including strain): Plasmodium vivax Etiologic Risk Group (see link below): http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0035406 */Disease Information (sort of like the Intro to your Mini Research Write up): Plasmodium vivax is a protozoal parasite and a human pathogen. The most frequent and widely distributed cause of recurring (Benign tertian) malaria, P. vivax is one of the six species of malaria parasites that commonly infect humans. It is less virulent than Plasmodium falciparum, the deadliest of the six, but vivax malaria can lead to severe disease and death due to splenomegaly (a pathologically enlarged spleen). It afflicted as many as eight U.S. presidents—including George Washington and Abraham Lincoln—and may have helped kill Genghis Khan. P. vivax is carried by the female Anopheles mosquito, since it is only the female of the species that bite. Link to TDR Targets page (if present): http://www.tdrtargets.org/targets/view?gene_id=269846 Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.) https://www.ncbi.nlm.nih.gov/protein/EDL46813.1 Essentiality of this protein: Is it a monomer or multimer as biological unit? (make prediction athttp://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Complex of proteins?: no Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): https://www.ebi.ac.uk/chembldb/index.php/target/inspect/10
*EC#: 2.7.7.6 Link to BRENDA EC# page: http://www.brenda-enzymes.org/enzyme.php?ecno=2.7.7.6 -- Show screenshot of BRENDA enzyme mechanism schematic Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): -- link to Sigma (or other company) page for assay (see Sigma links below) -- -or link (or citation) to paper that contains assay information http://www.ebi.ac.uk/chebi/searchId.do?chebiId=37416 -- links to assay reagents (substrates) pages. --- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model) -- PDB # or closest PDB entry if using homology model: 1RYQ -- For Homology Model option: ---- Show pairwise alignment of your BLASTP search in NCBI against the PDB ---- Query Coverage: 100% ---- Max % Identities: 141 ---- % Positives: n/a ---- Chain used for homology: 1RYQ_A
Current Inhibitors: 2-(3-(piperidin-4-yloxy)benzo[b]thiophen-2-yl)-5-((1,3,5-trimethyl-1H-pyrazol-4-yl)methyl)-1,3,4-oxadiazole (34c) Expression Information (has it been expressed in bacterial cells): n/a Purification Method: n/a Image of protein (PyMol with features delineated and shown separately): *Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
DYKFWYTQPVPKINDEFNESVNEPFISDNKVEDVRKDEYKLPPGYSWYVCDVKDEKDRSEIYTLLTDNYVEDDDNIFRFN
*length of your protein in Amino Acids: 384 residues Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 44890.56
Molar Extinction coefficient of your protein at 280 nm wavelength: 77490 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. *CDS Gene Sequence (paste as text only): gattataaattttggtatacccagccggtgccgaaaattaacgatgaatttaacgaaagc gtgaacgaaccgtttattagcgataacaaagtggaagatgtgcgcaaagatgaatataaa ctgccgccgggctatagctggtatgtgtgcgatgtgaaagatgaaaaagatcgcagcgaa atttataccctgctgaccgataactatgtggaagatgatgataacatttttcgctttaac tatagcgcggaatttctgctgtgggcgctgaccagcccgaactatctgaaaacctggcat attggcgtgaaatatgatgcgagcaacaaactgattggctttattagcgcgattccgacc gatatttgcattcataaacgcaccattaaaatggcggaagtgaactttctgtgcgtgcat aaaaccctgcgcagcaaacgcctggcgccggtgctgattaaagaaattacccgccgcatt aacctggaaaacatttggcaggcgatttataccgcgggcgtgtatctgccgaaaccggtg agcgatgcgcgctattatcatcgcagcattaacgtgaaaaaactgattgaaattggcttt agcagcctgaacagccgcctgaccatgagccgcgcgattaaactgtatcgcgtggaagat accctgaacattaaaaacatgcgcctgatgaaaaaaaaagatgtggaaggcgtgcataaa ctgctgggcagctatctggaacagtttaacctgtatgcggtgtttaccaaagaagaaatt gcgcattggtttctgccgattgaaaacgtgatttatacctatgtgaacgaagaaaacggc aaaattaaagatatgattagcttttatagcctgccgagccagattctgggcaacgataaa tatagcaccctgaacgcggcgtatagcttttataacgtgaccaccaccgcgacctttaaa cagctgatgcaggatgcgattctgctggcgaaacgcaacaactttgatgtgtttaacgcg ctggaagtgatgcagaacaaaagcgtgtttgaagatctgaaatttggcgaaggcgatggc agcctgaaatattatctgtataactggaaatgcgcgagctttgcgccggcgcatgtgggc attgtgctgctg *GC% Content for gene: 45.399306 *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): n/a *GC% Content for gene (codon optimized): n/a
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol) -- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
*NCBI Gene # or RefSeq#: 5473365
*Protein ID (NP or XP #) or Wolbachia#: XP_001614080.1
*Organism (including strain): Plasmodium vivax
Etiologic Risk Group (see link below): http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0035406
*/Disease Information (sort of like the Intro to your Mini Research Write up): Plasmodium vivax is a protozoal parasite and a human pathogen. The most frequent and widely distributed cause of recurring (Benign tertian) malaria, P. vivax is one of the six species of malaria parasites that commonly infect humans. It is less virulent than Plasmodium falciparum, the deadliest of the six, but vivax malaria can lead to severe disease and death due to splenomegaly (a pathologically enlarged spleen). It afflicted as many as eight U.S. presidents—including George Washington and Abraham Lincoln—and may have helped kill Genghis Khan. P. vivax is carried by the female Anopheles mosquito, since it is only the female of the species that bite.
Link to TDR Targets page (if present): http://www.tdrtargets.org/targets/view?gene_id=269846
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.) https://www.ncbi.nlm.nih.gov/protein/EDL46813.1
Essentiality of this protein:
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html):
Complex of proteins?: no
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): https://www.ebi.ac.uk/chembldb/index.php/target/inspect/10
*EC#: 2.7.7.6
Link to BRENDA EC# page: http://www.brenda-enzymes.org/enzyme.php?ecno=2.7.7.6
-- Show screenshot of BRENDA enzyme mechanism schematic
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
-- link to Sigma (or other company) page for assay (see Sigma links below)
-- -or link (or citation) to paper that contains assay information http://www.ebi.ac.uk/chebi/searchId.do?chebiId=37416
-- links to assay reagents (substrates) pages.
--- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 1RYQ
-- For Homology Model option:
---- Show pairwise alignment of your BLASTP search in NCBI against the PDB
---- Query Coverage: 100%
---- Max % Identities: 141
---- % Positives: n/a
---- Chain used for homology: 1RYQ_A
Current Inhibitors: 2-(3-(piperidin-4-yloxy)benzo[b]thiophen-2-yl)-5-((1,3,5-trimethyl-1H-pyrazol-4-yl)methyl)-1,3,4-oxadiazole (34c)
Expression Information (has it been expressed in bacterial cells): n/a
Purification Method: n/a
Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
DYKFWYTQPVPKINDEFNESVNEPFISDNKVEDVRKDEYKLPPGYSWYVCDVKDEKDRSEIYTLLTDNYVEDDDNIFRFN
YSAEFLLWALTSPNYLKTWHIGVKYDASNKLIGFISAIPTDICIHKRTIKMAEVNFLCVHKTLRSKRLAPVLIKEITRRI
NLENIWQAIYTAGVYLPKPVSDARYYHRSINVKKLIEIGFSSLNSRLTMSRAIKLYRVEDTLNIKNMRLMKKKDVEGVHK
LLGSYLEQFNLYAVFTKEEIAHWFLPIENVIYTYVNEENGKIKDMISFYSLPSQILGNDKYSTLNAAYSFYNVTTTATFK
QLMQDAILLAKRNNFDVFNALEVMQNKSVFEDLKFGEGDGSLKYYLYNWKCASFAPAHVGIVLL
*length of your protein in Amino Acids: 384 residues
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 44890.56
Molar Extinction coefficient of your protein at 280 nm wavelength: 77490
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):
gattataaattttggtatacccagccggtgccgaaaattaacgatgaatttaacgaaagc gtgaacgaaccgtttattagcgataacaaagtggaagatgtgcgcaaagatgaatataaa ctgccgccgggctatagctggtatgtgtgcgatgtgaaagatgaaaaagatcgcagcgaa atttataccctgctgaccgataactatgtggaagatgatgataacatttttcgctttaac tatagcgcggaatttctgctgtgggcgctgaccagcccgaactatctgaaaacctggcat attggcgtgaaatatgatgcgagcaacaaactgattggctttattagcgcgattccgacc gatatttgcattcataaacgcaccattaaaatggcggaagtgaactttctgtgcgtgcat aaaaccctgcgcagcaaacgcctggcgccggtgctgattaaagaaattacccgccgcatt aacctggaaaacatttggcaggcgatttataccgcgggcgtgtatctgccgaaaccggtg agcgatgcgcgctattatcatcgcagcattaacgtgaaaaaactgattgaaattggcttt agcagcctgaacagccgcctgaccatgagccgcgcgattaaactgtatcgcgtggaagat accctgaacattaaaaacatgcgcctgatgaaaaaaaaagatgtggaaggcgtgcataaa ctgctgggcagctatctggaacagtttaacctgtatgcggtgtttaccaaagaagaaatt gcgcattggtttctgccgattgaaaacgtgatttatacctatgtgaacgaagaaaacggc aaaattaaagatatgattagcttttatagcctgccgagccagattctgggcaacgataaa tatagcaccctgaacgcggcgtatagcttttataacgtgaccaccaccgcgacctttaaa cagctgatgcaggatgcgattctgctggcgaaacgcaacaactttgatgtgtttaacgcg ctggaagtgatgcagaacaaaagcgtgtttgaagatctgaaatttggcgaaggcgatggc agcctgaaatattatctgtataactggaaatgcgcgagctttgcgccggcgcatgtgggc attgtgctgctg
*GC% Content for gene: 45.399306
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): n/a
*GC% Content for gene (codon optimized): n/a
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):